E-2078, a potent, selective and stable dynorphin analog with preferential activity for the kappa-opioid receptor subtype on the mouse vas deferens neuroeffector junction

The profile of opioid activity of E-2078, a synthetic stable dynorphin analog, was examined in the mouse vas deferens bioassay and compared to that of methionine enkephalin and nonpeptide kappa agonists in the absence and in the presence of selective antagonists for the mu-, kappa- and delta-opioid receptor subtypes. The inhibitory action of E-2078 and related kappa agonists was specifically and potently antagonized only by norbinaltorphimine, revealing the presence of kappa receptors in this tissue and the predominant kappa activity of E-2078.     

Proteomic identification of mammalian cell surface derived glycosylphosphatidylinositol‐anchored proteins through selective glycan enrichment

Glycosylphosphatidylinositol‐anchored proteins (GPI‐APs) are an important class of glycoproteins that are tethered to the surface of mammalian cells via the lipid GPI. GPI‐APs have been implicated in many important cellular functions including cell adhesion, cell signaling, and immune regulation. Proteomic identification of mammalian GPI‐APs en masse has been limited technically by poor sensitivity for these low abundance proteins and the use of methods that destroy cell integrity. Here, we present methodology that permits identification of GPI‐APs liberated directly from the surface of intact mammalian cells through exploitation of their appended glycans to enrich for these proteins ahead of LC‐MS/MS analyses. We validate our approach in HeLa cells, identifying a greater number of GPI‐APs from intact cells than has been previously identified from isolated HeLa membranes and a lipid raft preparation. We further apply our approach to define the cohort of endogenous GPI‐APs that populate the distinct apical and basolateral membrane surfaces of polarized epithelial cell monolayers. Our approach provides a new method to achieve greater sensitivity in the identification of low abundance GPI‐APs from the surface of live cells and the nondestructive nature of the method provides new opportunities for the temporal or spatial analysis of cellular GPI‐AP expression and dynamics.     

The role of the Rieske iron-sulfur center as the electron donor to ferricytochrome c2 in Rhodopseudomonas sphaeroides

The Rieske iron-sulfur center in the photosynthetic bacterium Rhodopseudomonas sphaeroides appears to be the direct electron donor to ferricytochrome c₂, reducing the cytochrome on a submillisecond timescale which is slower than the rapid phase of cytochrome oxidation ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{3–5 μ}\text{s}$). The reduction of the ferricytochrome by the Rieske center is inhibited by 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole (UHDBT) but not by antimycin. The slower (1–2 ms) antimycin-sensitive phase of ferricytochrome c₂ reduction, attributed to a specific ubiquinone-10 molecule (Q_z), and the associated carotenoid spectral response to membrane potential formation are also inhibited by UHDBT. Since the light-induced oxidation of the Rieske center is only observed in the presence of antimycin, it seems likely that the reduced form of Q_z (Q_zH₂) reduces the Rieske center in an antimycin-sensitive reaction. From the extent of the UHDBT-sensitive ferricytochrome c₂ reduction we estimate that there are 0.7 Rieske iron-sulfur centers per reaction center.UHDBT shifts the EPR derivative absorption spectrum of the Rieske center from g_y 1.90 to g_y 1.89, and shifts the E_m,7 from 280 to 350 mV. While this latter shift may account for the subsequent failure of the iron-sulfur center to reduce ferricytochrome c₂, it is not clear how this can explain the other effects of the inhibitor, such as the prevention of cytochrome b reduction and the elimination of the uptake of H⁺_II; these may reflect additional sites of action of the inhibitor.     

A technique for the continuous recording of tick feeding electrograms and temperature by telemetry from free-ranging cattle

A biotelemetric technique is described for the continuous and simultaneous transmission of an analog signal of the feeding electrogram and of the temperature of a Hyalomma dromedarii female tick on a heifer roaming freely within a confined space beside an observation and control room. The tick was completely exposed to natural micrometeorological conditions. Circuit diagrams of a 102 MHz tick feeding electrogram transmitter and a 90 MHz tick temperature transmitter are included together with new methods used to hold and protect the instruments on the heifer.